Patterned, high surface area substrate with hydrophilic/hydrophobic contrast, and method of use

ABSTRACT

Nanoporous structures are constructed that have hydrophilic regions separated by hydrophobic regions. The porous, hydrophilic regions have reaction sites suitable for use in a bioassay application and have a higher density of reaction sites than that of a non-porous (2-D) surface. The structure may be made by depositing a layer of a matrix material (e.g., an organosilicate) and a porogen, and then crosslinking the matrix material to form a nanohybrid composite structure. The porogen is decomposed to form pores within the matrix material, and a reactive gas phase species (e.g., ozone) is patternwise directed onto a surface of the matrix material. Ultraviolet light (directed through a mask) activates the gas phase species to form a reactive species that then reacts with the matrix material to make it hydrophilic. The porogen may be decomposed thermally or by exposing it to an oxidizing atmosphere in the presence of ultraviolet light.

This application is a divisional of, and claims priority to, Applicant'sco-pending application Ser. No. 10/421,161 filed Apr. 22, 2003 andentitled “Patterned, high surface area substrate withhydrophilic/hydrophobic contrast, and method of use”, which is herebyincorporated by reference.

TECHNICAL FIELD

The invention relates to a process of forming arrays patterned intoregions of varying hydrophilicity, especially biomolecular arrays thathave high areal density and offer high signal to noise ratios.

BACKGROUND

Biomolecular arrays have quickly developed into an important tool inlife science research. Microarrays, or densely-packed, orderedarrangements of miniature reaction sites on a suitable substrate, enablethe rapid evaluation of complex biomolecular interactions. Because oftheir high-throughput characteristics and low-volume reagent and samplerequirements, microarrays are now commonly used in gene expressionstudies, and they are finding their way into significant emerging areassuch as proteomics and diagnostics.

The reaction sites of the array can be produced by transferring to thesubstrate droplets containing biological or biochemical material. Avariety of techniques can be used, including contact spotting,non-contact spotting, and dispensing. With contact spotting, a fluidbearing pin leaves a drop on the surface when the pin is forced tocontact the substrate. With non-contact spotting, a drop is pulled fromits source when the drop touches the substrate. With dispensing, a dropis delivered to the substrate from a distance, similar to an inkjetprinter. Reaction sites on the array can also be produced byphotolithographic techniques (such as those employed by Affymetrix orNimbleGen, for example).

The quality of the reaction sites directly affects the reliability ofthe resultant data. Ideally, each site would have a consistent anduniform morphology and would be non-interacting with adjacent sites, sothat when a reaction occurred at a given site, a clear and detectableresponse would emanate from only that one site, and not from neighboringsites or from the substrate. To reduce the overall size of an arraywhile maximizing the number of reaction sites and minimizing therequired reagent and sample volumes, the sites on the array should havethe highest possible areal density.

With current microarray technology, which is dominated by the use offlat substrates (often glass microscope slides), areal density islimited. To increase the signal from a given reaction site, theinteraction area between the fluid (usually aqueous) and the substrateshould be maximized. One way to do this is by using a surface thatpromotes wetting. A flat surface that promotes wetting, however, canlead to spots (and thus reaction sites) having irregular shapes andcompositions. A flat wetting surface can also lead to the spreading offluid from its intended site into neighboring sites. Thus, flat surfacesare intrinsically limited by fluid-surface interactions that force atradeoff between the desired properties of the reaction sites.

To make the sites more uniform, the surface can be made non-wetting.Unfortunately, this reduces the interaction area between the fluid andthe surface, thereby reducing the signal that would otherwise beobtainable. In addition, since droplets do not adhere well to a flatnon-wetting surface, deposition volumes can vary from site to site, anddroplets can slide away from their intended location, unless they areotherwise confined.

One way of avoiding the wetting vs. non-wetting dichotomy is to preparesurfaces that have regions of varying hydrophilic/hydrophobic contrast.Due to the aqueous environment of biomolecular arrays, patterned mediahaving hydrophilic/hydrophobic contrast are ideal for confiningbioactivity to within discrete regions defined by the pattern, with eachdiscrete region in effect acting as an individual bio-probe. Ahydrophobic surface is generally regarded as one having a static watercontact angle of greater than 90 degrees, with decreasing contact anglesresulting in progressively more hydrophilic surfaces. A surface having awater contact angle of less than 65 degrees is considered stronglyhydrophilic. (For a discussion of contact angles, see A. W. Adamson etal., “Physical chemistry of surfaces”, John and Wiley & Sons, New York,1997.)

Several methods have been reported for preparing patterns of varyinghydrophilicity, including traditional lithographic methods, imprinting,and contact printing. Lithographic techniques rely on the attachment ofhydrophobic (or hydrophilic) molecules to preselected regions defined byphotoresists in a hydrophilic (or hydrophobic) matrix. (See, forexample, J. H. Butler et al., J. Am. Chem. Soc. 2001, 123, 8887.) Withimprinting techniques, hydrophilic regions are created by pipettingdroplets of a washable or hydrophilic lacquer, much like that in anink-jet printer, and then converting the adjacent regions to hydrophobicregions. (See, for example, UK Patent Application GB 2340298AUK andPatent Application GB 2332273A.) Contact printing methods typicallyinvolve elastomeric stamps with hydrophilic (or hydrophobic) inks, withhydrophilic (or hydrophobic) patterns being generated as a result oftransferring the ink onto a substrate. (See, for example, G. MacBeath etal, Science 2000, 289, 1760; and C. M. Niemeyer et al., Angew. Chem.Int. Ed. 1999, 38, 2865). U.S. Pat. No. 5,939,314 to Koontz disclosesporous polymeric membranes having hydrophilic/hydrophobic contrast, inwhich the pore size is on the order of 0.1-2000 microns, but pores ofthis size are still relatively large. These methods generally involve,however, a series of several process steps.

A simple, more effective route to patterned substrate arrays havingregions of varying hydrophilic/hydrophobic contrast would be highlydesirable. Further, such arrays should have a high areal density ofsites and high effective surface area to permit the collection of datawith good signal/noise ratio. In addition, such an apparatus wouldideally have sites of consistent and uniform spot morphology.

SUMMARY OF THE INVENTION

A simple and effective method is disclosed for generating films thatinclude 3-D, nanoporous hydrophilic regions separated by hydrophobicregions. The porous, hydrophilic regions have reaction sites suitablefor receiving reagents and/or reactants (biological, biochemical, orotherwise) that can be detected when tagged with a compound thatfluoresces in response to irradiation with light (UV light, forexample). The emitted fluorescence can then be detected by an opticaldetector. An advantage of porous material is that the density ofpotential reaction and/or absorption sites is significantly higher thanthat provided by a non-porous (2-D) surface. Patterning of the substratemay be accomplished by directing ultraviolet light onto a mask in thepresence of a latent oxidizing species, such as ozone. Alternatively, anO₂-RIE process or oxygen plasma may be used in conjunction with a shadowmask to pattern the film.

An advantage of preferred methods disclosed herein is that the porosityof the films may be controlled by incorporating a pore-generating agentor compound (porogen) into a host material, followed by decomposition ofthe porogen. By utilizing porogen compounds in this manner, pore sizesand porosity can be tailored to the user's needs. One advantage of theUV/ozone treatments disclosed herein is that they are an economical wayof producing reactive oxidizing species that can be utilized to produceregions of hydrophilic/hydrophobic contrast. Another advantage of theUV/ozone treatments is that the feature resolution (i.e., the spacingbetween adjacent hydrophobic and hydrophilic features) can be controlledoptically.

One preferred implementation of the invention is a method of formingdiscrete hydrophilic regions on a substrate. The method includesdepositing a layer onto a substrate, in which the layer includes amatrix material and a porogen, and then crosslinking the matrix materialto form a nanohybrid composite structure out of the matrix material andthe porogen. The method also includes decomposing the porogen to formpores within the matrix material, and patternwise directing a reactivegas phase species onto a surface of the matrix material. This formsdiscrete regions in the matrix material that are more hydrophilic thanare other regions in the material that are adjacent to these discreteregions. The discrete (relatively more hydrophilic) regions and theother (preferably hydrophobic) regions extend from the surface of thematerial to beneath the surface of the material. The matrix material ispreferably an organosilicate material, in which the nanohybrid structureis formed by crosslinking the organosilicate material. This crosslinkingmay induce phase separation between the organosilicate material and theporogen, or if a templating approach is used, the crosslinking of theorganosilicate material is templated by the porogen. The crosslinkingmay be thermally induced, or it may be induced through a photochemicalprocess, e-beam irradiation, or the addition of a basic or acidiccatalyst to the organosilicate material. The gas phase species isadvantageously directed onto the organosilicate material in apatternwise way, so that the discrete regions form a pattern within theorganosilicate material. This pattern may be formed by a mask inproximity with the layer, with less hydrophilic regions in theorganosilicate material corresponding to opaque portions of the mask.The gas phase species preferably includes an oxidizing species, such asozone, with ultraviolet radiation activating the gas phase species toform a reactive species. The decomposition of the porogen may bethermally induced, or alternatively, this decomposition may be inducedby exposing the porogen to an oxidizing atmosphere in the presence ofelectromagnetic radiation. The dimensions of the relatively morehydrophilic regions may be selected to be suitable for use in abiomolecular array. Because the pores are suitable for concentratingreagents and reactants (like those in a bioassay application), thedimensions of these hydrophilic regions may be chosen to be smaller thanthey otherwise would be in the absence of the pores.

One preferred implementation of the invention is a method of formingregions of varying hydrophilicity on a substrate. The method includesdepositing a layer onto a substrate, in which the layer includes amatrix material and a porogen, and then crosslinking the matrix materialto form a nanohybrid composite structure out of the matrix material andthe porogen. The method further includes thermally decomposing theporogen to form pores in the layer, and patternwise oxidizing the matrixmaterial to form regions of varying hydrophilicity within the layer,with these regions extending from a surface of the material into thematerial itself (e.g., 10 microns or less). This patternwise oxidizingmay include directing, in the presence of a gas phase species such asozone, ultraviolet radiation through a mask that is in proximity withthe layer. The regions preferably include discrete hydrophilic regionsseparated by hydrophobic regions.

Another preferred implementation of the invention is a method of formingregions of varying hydrophilicity on a substrate. The method includesdepositing a layer onto a substrate, in which the layer includes amatrix material and a porogen. The method further includes crosslinkingthe matrix material to form a nanohybrid composite structure out of thematrix material and the porogen, and patternwise directing (in thepresence of an oxidizing species such as ozone) ultraviolet radiationonto selected regions of the matrix material to both decompose theporogen and induce hydrophilicity within the selected regions, withthese selected regions extending from a surface of the matrix materialinto the material. The matrix material is preferably an organosilicate,with the crosslinking including thermally crosslinking the matrixmaterial. The selected regions are preferably hydrophilic regionssurrounded by hydrophobic regions.

One preferred embodiment is a nanoporous structure having regions ofvarying hydrophilicity, in which the regions correspond to a preselectedpattern. The structure includes pores having a minimum characteristicdimension of between 2 nm and 75 nm, with these pores constituting atleast 5% of the structure by volume. These regions of varyinghydrophilicity preferably include discrete hydrophilic regions separatedfrom each other by hydrophobic regions. The thickness of these regionsmay be less than 10 microns, e.g., between 0.5 and 10 microns, with theregions having a characteristic dimension between 2 microns and 1000microns. In another preferred embodiment, the pores constitute at least30% of the structure by volume.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1, which includes FIGS. 1A, 1B, 1C, 1D, 1E, 1F, and 1G, illustratessteps that may be used in forming a layer that includes porous,hydrophilic regions surrounded by hydrophobic regions, in which thesequence of steps represented by FIGS. 1A, 1B, 1C, 1D, and 1E representsone preferred method, and the sequence of steps represented by FIGS. 1A,1B, 1F, and 1G represents another preferred method.

FIG. 2 is a schematic illustration of how functional groups inpolymethylsilsesquioxane (PMSSQ) are modified as a result of exposure toultraviolet light and ozone.

FIG. 3 illustrates the effect of temperature and exposure time on thestatic water contact angle of a layer of porous PMSSQ when the layer isexposed to ultraviolet light and ozone.

FIG. 4 is an image of drops of water on a 1″ diameter layer of porousPMSSQ that has been patterned into hydrophobic and hydrophilic regions.

FIG. 5 illustrates a fluorescent dye structure attached to a linker thatin turn was attached to a layer of porous PMSSQ that had been subjectedto an ultraviolet light/ozone treatment.

FIG. 6 is a fluorescence microscope image of a porous organosilicatesurface that has been patterned into hydrophobic and hydrophilicregions, in which the hydrophilic regions have been tagged with thefluorescent dye of FIG. 5.

FIGS. 7A, 7B, and 7C are fluorescence microscope images of porous PMSSQpatterned into hydrophobic and hydrophilic regions, in which thesmallest characteristic feature sizes (the line widths of the segmentsin the images) are 32, 16, and 8 micrometers, respectively.

FIG. 8 shows how the refractive index of a nanohybrid composite filmchanges as a function of UV/ozone treatment time at temperature of 30°C.

DETAILED DESCRIPTION OF THE INVENTION

Methods are disclosed herein for generating 3-D, nanoporous structureshaving regions of varying hydrophilic/hydrophobic contrast, e.g.,alternating hydrophilic and hydrophobic regions. In one preferredmethod, a patterned nanoporous organosilicate is formed by first formingpores within a layer and then patterning the porous layer into regionsof varying hydrophilicity. In another preferred method, a single processstep is employed to make preselected regions of a substrate both porousand relatively hydrophilic with respect to adjacent regions in thesubstrate.

FIG. 1A shows a substrate 20 onto which a solution is applied. Thesubstrate may be silicon, silicon dioxide, fused glass, ceramic, metal,or any other suitable material. The solution preferably includes a hostmatrix material (such as an organosilicate) and a decomposable porogendissolved in a suitable solvent (e.g., 1-methoxy-2-propanol acetate).The porogen may be chemically bonded to the matrix material eitherdirectly or through a coupling agent, as discussed in U.S. Pat. No.6,107,357 issued Aug. 22, 2000 to Hawker et al., which is herebyincorporated by reference. The solution may be applied to the substrate20 by spraying, spin coating, dip coating, or doctor blading, so that auniform thin film 26 of a porogen/matrix material mixture remains on thesubstrate 20 after the solvent has evaporated. Preferred matrixmaterials include organosilicates, such as those disclosed in U.S. Pat.No. 5,895,263 issued Apr. 20, 1999 to Carter et al. (which is herebyincorporated by reference), including the family of organosilicatesknown as silsesquioxanes, (RSiO_(1.5))_(n). Suitable silsesquioxanes forthe present invention include hydrido (R═H), alkyl (R=methyl), aryl(R=phenyl) or alkyl/aryl, as well as polymethylsilsesquioxane (PMSSQ),which are commercially available from Dow Corning, Techneglas, LGChemicals, and Shin-Etsu, for example. Other suitable matrix materialsinclude polysilanes, polygermanes, carbosilanes, borozoles, carboranes,the refractory oxides, amorphous silicon carbide, and carbon dopedoxides. Suitable decomposable porogens include linear polymers,crosslinked polymeric nanoparticles, block copolymers, randomcopolymers, dendritic polymers, star polymers, hyperbranched polymers,grafts, combs, unimolecular polymeric amphiphiles, and porogens such asthose discussed in U.S. Pat. No. 5,895,263 to Carter et al. Asillustrated in FIG. 1B, a nanohybrid composite structure between theporogen 32 and the matrix 38 is then formed, so that the porogen isentrapped in the crosslinked matrix. Different processes may be employedto arrive at this stage, such as i) a nucleation and growth process andii) a particle templating process. In a nucleation and growth process,the sacrificial porogen is miscible in the matrix material before curingand phase separates upon the crosslinking of the matrix material to formpolymer-rich domains. (Crosslinking is preferably accomplished byheating the matrix material, although other ways of initiatingcrosslinking are possible, such as photochemical means, e-beamirradiation, and the addition of a basic or acidic catalyst to theorganosilicate material.) Ideally, the domains remain nanoscopic due tolow mobility in the viscous, crosslinking matrix, and these domainsultimately become the pores. The morphology and size of the poresdepends on the loading level of the porogen (i.e., how much porogen ispresent in the matrix prior to decomposition of the porogen), theporogen molecular weight and structure, resin structure, processingconditions, and so on. Although small pores can be generated, theprocess has many variables.

In a porogen templating process, on the other hand, the porogen is neverreally miscible in the matrix, but is instead dispersed. The matrixcrosslinks around the porogen, so that the porogen templates thecrosslinked matrix. (Below the percolation threshold, the porousmorphology is composition independent, one porogen molecule generatesone hole, and pore size depends on the porogen size. Therefore, it isadvantageous to work above the percolation threshold, so thatinterconnected pores are formed.) Templating behavior is observed in theacid-catalyzed hydrolytic polymerization of tetraethoxysilane (TEOS) inthe presence of surfactant molecules (see R. D. Miller, Science, 1999,286, 421 and references cited therein). The surfactant molecules formdynamic supermolecular structures which upon processing template thecrosslinked matrix material. Templating behavior is often observed forhighly crosslinked nanoparticles generated by suspension (see M. Munzer,E. Trommsdorff, Polymerization in Suspension, Chapter 5 inPolymerization Processes, C. F. Schieldknecht, editor, WileyInterscience, New York, 1974) or emulsion polymerization (see D. H.Blakely, Emulsion Polymerization: Theory and Practice, Applied Science,London, 1965); these are classified as top down approaches to porogensynthesis. Bottom up approaches to crosslinked nanoparticles are alsopossible, and may involve the intramolecular crosslinking collapse of asingle polymer molecule to produce a crosslinked nanoparticle (see D.Mercerreyes et al., Adv. Mater. 2001, 13(3), 204; and E. Harth et al.,J. Am. Chem. Soc., 2002, 124, 8653). A bottom up templating approach mayalso be observed for un- or lightly-crosslinked materials which exhibitparticle-like behavior in the matrix, e.g., with multiarm star-shapedpolymeric amphiphiles where the core and shell portions have widelydifferent polarity. In this case, the inner core collapses in the matrixmaterial while the polymer corona stabilizes the dispersion to preventaggregation (see U.S. Pat. No. 6,399,666 issued Jun. 4, 2002 to Hawkeret al., which is hereby incorporated by reference). Each of theseporogen classes (surfactant, top down, and bottom up) may be used totemplate the crosslinking of, for example, PMSSQ.

Thus, more than one approach may be used to generate the porogen phase32 within the matrix 38 shown in FIG. 1B. For systems displayingnucleation and growth characteristics, the matrix 38 (e.g., theorganosilicate) and the porogen 32 are subjected to a phase separationprocess. A preferred way of inducing this phase separation is by heatingthe (preferably thin, <5 microns) film 26 to the crosslinking reactiontemperature of the organosilicate, thereby forming a nanohybridcomposite of the porogen and organosilicate in the film, so that anorganic, porogen phase 32 is entrapped in an inorganic, crosslinkedmatrix 38. Alternatively, a templating approach may be used, asdiscussed above, in which a suitable porogen 32 is dispersed but is notmiscible in an appropriate matrix 38, which is then thermoset (uponapplication of heat, for example) to form a nanohybrid structure.Regardless of which approach is used (nucleation/growth or templating),the loading level of the porogen is preferably high enough that thepercolation threshold is reached in the nanohybrid composite and porousfilm so derived, so that the pores 44 are highly interconnected (notshown in the cross sectional views of FIG. 1). When the pores 44 areinterconnected in this manner, the effective surface area of the endproduct (corresponding to FIG. 1E or 1G) is high, and theinterconnectivity of the pores facilitates accessibility to reactantsand reagents. This permits good signal/noise ratio data in abiodetection application. To this end, a porogen loading of 30 wt. % ormore is preferred, resulting in an end product whose volumetric porosityis approximately 30%.

At this point, more than one approach may be employed to produce ananoporous structure having regions of varying hydrophilic/hydrophobiccontrast, as indicated by the two pathways corresponding to FIGS. 1C and1F, respectively. Either of these pathways, however, may be used togenerate interconnected pores that preferably have an averagecharacteristic minimum dimension (e.g., a diameter) of between 2 nm and75 nm, and still more preferably between 2 nm and 50 nm. Pores of thissize are advantageous in that they offer the user high effective surfacearea and access to reagents and reactants. In FIG. 1C, additional heatis applied to the film to bring it to a temperature above thedecomposition temperature of the porogen, e.g., the film may be heatedto 350° C. or above in an inert atmosphere. This results in the thermaldecomposition of the phase-separated porogen 32, so that the spaceoccupied by the porogen becomes voids 44 or pores. This approach to thegeneration of a nanoporous film, known as the sacrificial porogen (poregenerator) approach, relies on the selective removal of the organicmacromolecular (porogen) phase from phase-separated mixtures of organic(or inorganic) polymers. (Further details on porogens may be found inU.S. Pat. No. 5,895,263 to Carter et al., for example.) The morphologyand dimensions of the pores 44 are determined mainly by the interactionbetween the porogen (the dispersed phase 32), the organosilicate matrix38, and the composition of these mixtures. In general, with increasingporogen loading level (i.e., increasing weight percentage of the porogenin the organosilicate prior to decomposition of the porogen), the poresformed in the organosilicate become increasingly interconnected: For lowporogen loading (<20%), a closed cell structure is observed, whereas forhigher porogen loading, interconnected or bicontinuous phase structuresare observed. Using the methods described herein, end products may beobtained whose volumetric fraction of pores is between 5% and 80%, andmore preferably between 30% and 70%.

The film may then be exposed to ultraviolet (UV) light in the presenceof ozone (O₃), as indicated by the arrows 48 of FIG. 1D, to generateregions of varying hydrophilicity. By patternwise exposing the filmthrough use of a mask 50, regions of the film that are so exposed becomerelatively more hydrophilic regions 60, as shown in FIG. 1E. As analternative to the UV/ozone process (in which O₃ is photodissociated byUV light to generate atomic oxygen, which is a reactive species), aUV/N₂O process (in which N₂O is photodissociated by UV light to generateatomic oxygen) or a UV/H₂O₂ process (in which H₂O₂ is photodissociatedby UV light to generate the hydroxyl radical, which is also a reactivespecies) may be used in conjunction with a mask 50. Other sources ofhydroxy, alkoxy, and aryloxy radicals may be used instead of H₂O₂, suchas RO₂H, RO₂R′, and RCO₃R′, in which R and R′ are alkyl or arylsubstituents.

The portions of the mask 50 shown as darkened regions represent opaqueportions 50 b of the mask, and the lighter regions represent portions 50a of the mask that are open spaces or at least transparent to UV light.(For example, if the portions 50 a are quartz, the mask 50 may belocated slightly above the film, with ozone being passed between themask and the film. Alternatively, the mask 50 may be placed in directcontact with the film, with ozone being diffused directly through theporous film.) On the other hand, those regions 64 of the film thatremain unexposed to UV, and therefore unexposed to reactive oxygen(i.e., those regions shielded by the opaque portions 50 b), remainhydrophobic. The mask 50 can be metallic (e.g., chromium, copper, brass,or beryllium-copper) and is positioned above the film, preferably indirect contact with the film, to facilitate good spatial contrastbetween the relatively hydrophilic regions 60 and the surroundinghydrophobic regions. Masks similar to those used in the photolithographyindustry may be employed, with a spatial resolution (the distancebetween the opaque portions 50 b and the open portions 50 a) being lessthan 1 micron, for example. As an alternative to the UV/ozone treatment,an oxidizing plasma (e.g., O₂) may be directed onto a shadow mask. Inanother implementation, an O₂—RIE process in combination with a shadowmask may be used to form the hydrophilic regions 60, or any direct-writeoxidizing source (e.g., an ion beam) may be used for this purpose.

The chemical mechanism leading to the desired hydrophilicity can be atleast partially explained as follows. Generally, it is known that ozoneis “activated” to produce a reactive species (atomic oxygen) uponabsorption of UV light (e.g., the 253.7 nm Hg line may be used tophotodissociate ozone). Atomic oxygen is postulated to be an etchingspecies, which, over a wide range of temperatures (e.g., from roomtemperature to ˜300° C. and higher), is capable of breaking organicmaterials into simple, volatile oxidation products such as carbondioxide, water, and so on. It is believed that the UV/ozone treatment(or alternatively, the UV/N₂O treatment or the UV/H₂O₂ treatmentdiscussed above) eliminates matrix methyl groups (—CH₃) from the PMSSQand introduces a polar oxidation product, namely hydroxyl groups (—OH),as shown in FIG. 2. FTIR spectroscopy measurements reveal that aprominent absorption band at 3400 cm⁻¹ arises as a result of theUV/ozone treatment, suggesting that hydroxyl groups are present in theUV/ozone treated sample. Thus, the silicon species left behind afteroxidation of PMSSQ contains a significant amount of polar SiOHfunctionality, which is known to be hydrophilic. Directing an oxidizingspecies onto other matrix materials, such as polysilanes, polygermanes,carbosilanes, borozoles, carboranes, the refractory oxides, amorphoussilicon carbide, and carbon doped oxides, also leads to the formation of—OH.

As an alternative to the series of steps illustrated by FIGS. 1C, 1D,and 1E, the steps illustrated by FIGS. 1F and 1G may be used after thephase separation of FIG. 1B. In FIG. 1F, a UV/ozone treatment incombination with a mask 50 is used. This technique generates porous,hydrophilic regions 60 separated from non-porous, hydrophobic regions 64a, as shown in FIG. 1G. In this case, the UV/ozone treatment decomposesthe organic, porogen phase 32 (into CO₂, H₂O, and lower molecular weightoxidized fragments) while simultaneously changing the chemical propertyof the organosilicate to produce hydrophilic regions 60. (For thisreason, the regions 50 a in the mask of this implementation arepreferably open spaces that allow the decomposing porogen to diffuse outof and away from the film.) This approach is advantageous in that fewerprocess steps are involved than the approach that includes the stepsillustrated by FIGS. 1C, 1D, and 1E. Furthermore, the step illustratedby FIG. 1F allows the user to control how far into the film pores 44 areformed by controlling the ozone concentration, ultraviolet lightintensity, temperature, and/or exposure time. Increasing any one ofthese three variables tends to form pores deeper into the film, andthereby tailor the volume available to the user, e.g., in a biodetectionexperiment.

The methods disclosed herein may be used to form porous films having athickness of up to at least 1 micron. Film thicknesses in the ranges of0.5-1 micron, 0.5-2 microns, 0.5-3 microns, 0.5-4 microns, 0.5-5microns, 0.5-10 microns or more may also be realized. In addition,well-defined feature sizes as small as about 4 microns may be obtained,as discussed in Example 4 below. Feature sizes in the ranges of 2-4microns, 2-10 microns, 2-50 microns, 2-1000 microns, 4-50 microns, 4-75microns, 4-500 microns, and 4-1000 microns may also be realized.

The hydrophilic/hydrophobic patterning techniques described herein maybe used to form 3-D porous structures or be applied to non-porousstructures yielding surfaces of hydrophilic/hydrophobic contrast. Forexample, the UV/ozone technique (and the UV/H₂O₂ and UV/N₂O techniques)may be applied to form (non-porous or nominally porous) surfaces thatare patterned into hydrophilic and hydrophobic regions. Such surfacescan be used in a biodetection application. Materials that may be used insuch a 2-D patterning technique (in addition to the matrix materialsalready described) include the family of silicon containing polymersthat are not silicates or silicones, as well as carbon-containingpolymers that do not contain silicon.

EXAMPLES

The porous PMSSQ of Examples 1-5 was formed by beginning with a mixtureof 80 wt. % porogen (namely, the triblock copolymer of ethylene oxideand propylene oxide sold under the name “Pluronics” by the BASF company)and 20 wt. % organosilicate (namely, the polymethylsilsesquioxane GR650Ffrom Techneglas, shown in FIG. 2) dissolved in the solvent1-methoxy-2-propanol acetate. This solution was applied uniformly to asilica wafer by spin coating, so that a uniform thin film of theporogen/organosilicate mixture remained on the substrate 20 after thesolvent had evaporated. A nanohybrid composite film was produced byheating the porogen/organosilicate mixture (at a temperature of between150° C. and 250° C.) in an inert atmosphere.

For Examples 1-4, porosity in the nanohybrid composite film was thengenerated by heating it to 350° C. or higher. The porous film was thensubjected to a UV/ozone treatment to generate regions of varyinghydrophilicity. For Example 5, a UV/ozone treatment was applied to thenanohybrid composite film at a temperature of 30° C., which generatedporosity in the film as well as regions of varying hydrophilicity.

The UV/ozone treatment for these examples was performed as follows. Theoxygen flow rate into the ozone generator was 3.0 standard liters permin, thereby producing an ozone concentration of 38000 ppm by volume.For this purpose, a SAMCO International, Inc. UV/ozone stripper (modelUV-300H) was used. The UV light source included two 235 watt hotcathodes, low-pressure, high-output mercury vapor lamps, having primaryprocess wavelengths at 254 nm and 185 nm.

Example 1

Static water contact angle measurements were made with an AST VideoContact Angle System 2500 XE to quantify the effect of UV/Ozonetreatment (like that shown in FIG. 1D) on the surface properties ofporous PMSSQ films (like that shown in FIG. 1C). FIG. 3 shows thecontact angle as a function of treatment time for porous film producedfrom starting material of 80 wt. % porogen/20 wt. % organosilicate.(Films of 10, 30, and 50 wt. % porogen were examined as well, and gavesubstantially similar results; films with a higher initial wt. % ofporogen have greater porosity following decomposition of the porogen.)There is a rapid decrease in the contact angle over time, indicatingthat the surface is becoming more hydrophilic. This phenomenon isaccelerated at higher temperatures, as a comparison between the data at30° C. and 150° C. shows. A still more rapid decrease in the contactangle was observed at 250° C. The water contact angle decreases frommore than 100 degrees initially to 10 degrees or less (see the 150° C.data, for example). The contact angle data of FIG. 3 are clear evidencethat the surface of the PMSSQ film becomes hydrophilic as a result ofthe UV/ozone treatment, and that the degree of this hydrophilicity canbe controlled (e.g., by controlling treatment time and temperature) overthe range from between 90 degrees down to about 10 degrees or less.

Example 2

By limiting UV exposure to those areas on a film corresponding to openareas within a metal mask (as shown by the mask of FIG. 1D, forexample), hydrophilic patterns in a hydrophobic matrix can be obtained.In this case, only those areas on the film exposed to both UV and ozonebecome hydrophilic, while unexposed areas remain hydrophobic. Masks orschemes which create patterns of UV light are useful for thispatterning. The result of such a patterning process is demonstrated inFIG. 4, which shows porous PMSSQ (on a 1″ silica wafer) on which waterdroplets are confined to ¼ inch diameter hydrophilic areas.

Example 3

When hydrophilic areas are reduced in size to the point that they have acharacteristic dimension (i.e., an approximate width or length) of 250microns or less, the surface tension of water prevents the formation ofwell-defined drops (like those shown in FIG. 4), so that only wavyshapes at the water/surface/air contact line are evident, indicatingthat probe molecules in aqueous solution can be confined to thehydrophilic patterned areas. Indeed, the surface hydroxyl groupsgenerated by UV/Ozone treatment are themselves useful for chemicalreactions for bonding probe molecules covalently.

To demonstrate that a higher number density of —OH groups is availablewithin a i) UV/ozone treated porous organosilicate medium than eitherii) a flat silica substrate that was not treated with UV/ozone or iii)non-porous MSSQ treated with UV/ozone, a fluorescent dye was used.Specifically, the linker 3-bis(2-hydroxyethyl) amino propyltriethoxysilane was attached to —OH groups on representative samples ofi), ii), and iii). The fluorescent dye 6-carboxyfluorescein(commercially available from Applied Biosystems as 6-FAM™ amidite, forexample) was then selectively attached to each of these samples, asindicated in FIG. 5. This dye fluoresces green in response to opticalexcitation.

FIG. 6 shows a fluorescence microscope image of a porous, patternedsurface (case i) to which the linker and fluorescent dye have beenattached. Images were obtained using a fluorescence microscope, and theintensity of the fluorescent image was quantified using image analysissoftware. The image of FIG. 6 shows discrete regions where the dye hasbeen selectively attached, with these regions corresponding to thepatterned areas where surface SiOH functional groups have beengenerated. These discrete regions, which are clearly contrasted from theunderlying matrix, are roughly circular and have a diameter ofapproximately 250 μm.

Continuing with this example, the fluorescence intensity (of greenlight) from these discrete, circularly shaped regions was compared withthat from samples ii) and iii). The use of image analysis softwaresuggests that the signal intensity was approximately 10 times highersignal intensity from porous PMSSQ surface (case i) than from a nativeoxide layer of a flat silicon wafer that was not treated by UV/ozone(case ii), and about 7 times higher than the signal from a non-porousPMSSQ surface exposed to the same UV/ozone treatment (case iii). Theenhanced patterned fluorescence of the treated PMSSQ surface relative tonative oxide shows that 2-D images can be produced in denseorganosilicate films using the technique. The quantitative data areclear evidence of a volumetric effect, namely, that porous PMSSQsurfaces allow for a greater number density of attached molecules thando their non-porous counterparts, indicating that —OH groups are formedthroughout the porous sample.

Example 4

Photolithographic masks (of quartz and a chromium coating) havingdifferent features sizes were placed in direct contact with 750 nm thickporous PMSSQ film to make hydrophilic/hydrophobic patterns correspondingto the features of the masks. Fluorescent dye was attached tohydrophilic regions of the porous PMSSQ film, in a manner like thatdescribed above in connection with Example 3. FIGS. 7A, 7B, and 7C showdarker (hydrophobic) regions and lighter, fluorescing (hydrophilic)regions, in which fluorescent dye has been attached to the hydrophilicregions. FIGS. 7A, B, and C show well defined patterns of 32, 16, and 8μm feature sizes, respectively (corresponding to the width of the darksegments in these figures). For features sizes smaller than 4 μm, therewas some evidence of smeared boundaries between the hydrophilic andhydrophobic regions, presumably due to diffusion of the active oxidizerbefore reaction with the matrix.

Example 5

The refractive index of a nanohybrid composite film was measured toquantify porogen decomposition as a function of UV/ozone treatment time.The temperature was held constant at 30° C. A white light interferometer(Filmetrics F20 Thin Film Measurement System) was used to measure therefractive index. FIG. 8 shows how the refractive index changes as afunction of UV/ozone treatment time. Prior to any UV/ozone treatment(time=0 minutes), the nanohybrid composite film has a refractive indexof 1.44. The refractive index decreases as the UV/ozone treatment isapplied. This is attributed to decomposition of the porogen, leading toan increased volumetric fraction of air within the film. The refractiveindex reaches about 1.20 after 40 minutes of this treatment, which isvery nearly equal to the index of refraction of a porous film whoseporosity has been generated by thermal decomposition of the porogen.

The invention may be embodied in other specific forms without departingfrom its spirit or essential characteristics. The described embodimentsare to be considered in all respects only as illustrative and notrestrictive. The scope of the invention is therefore indicated by theappended claims rather than the foregoing description. All changeswithin the meaning and range of equivalency of the claims are to beembraced within that scope.

1. A structure formed from material that has been patterned into regionsof varying hydrophilicity, wherein the regions correspond to apreselected pattern, the structure includes pores having a diameter ofbetween 2 nm and 75 nm, and the structure has a volumetric porosity ofat least 5%.
 2. The structure of claim 1, wherein the regions of varyinghydrophilicity include discrete hydrophilic regions separated from eachother by hydrophobic regions.
 3. The structure of claim 2, wherein thethickness of the structure is less than 10 microns.
 4. The structure ofclaim 2, wherein the thickness of the structure is between 0.5 and 10microns.
 5. The structure of claim 2, wherein the regions have acharacteristic dimension between 2 microns and 1000 microns.
 6. Thestructure of claim 2, wherein the structure has a volumetric porosity ofat least 30%.
 7. The structure of claim 2, wherein the regions have acharacteristic dimension between 4 microns and 500 microns.
 8. Thestructure of claim 2, wherein the regions have a characteristicdimension between 4 microns and 75 microns.
 9. The structure of claim 2,wherein the regions have a characteristic dimension between 2 micronsand 50 microns.
 10. The structure of claim 2, wherein the thickness ofthe structure is at least one micron.
 11. The structure of claim 2,further comprising a substrate on which the structure is formed.
 12. Thestructure of claim 1, comprising porous, hydrophilic regions oforganosilicate material separated from non-porous, hydrophobic regionsof organosilicate material.
 13. The structure of claim 1, comprisingporous, hydrophilic regions of organosilicate material separated fromporous, hydrophobic regions of organosilicate material.
 14. A structureformed from material that has been patterned into regions of varyinghydrophilicity, wherein the regions correspond to a preselected pattern,the structure includes pores having a diameter of between 2 nm and 50nm, and the structure has a volumetric porosity of at least 5%.
 15. Thestructure of claim 14, wherein the regions of varying hydrophilicityinclude discrete hydrophilic regions of organosilicate materialseparated from each other by hydrophobic regions of organosilicatematerial.
 16. The structure of claim 15, wherein the thickness of thestructure is between 0.5 and 10 microns.
 17. The structure of claim 15,wherein the regions have a characteristic dimension between 2 micronsand 1000 microns.
 18. The structure of claim 15, wherein the structurehas a volumetric porosity of at least 30%.
 19. The structure of claim15, wherein the regions have a characteristic dimension between 2microns and 50 microns.
 20. The structure of claim 15, wherein thethickness of the structure is at least one micron.
 21. The structure ofclaim 15, further comprising a substrate on which the structure isformed.
 22. The structure of claim 14, comprising porous, hydrophilicregions separated from non-porous, hydrophobic regions.
 23. Thestructure of claim 14, comprising porous, hydrophilic regions separatedfrom porous, hydrophobic regions.